An estimated 150 million UTIs occur annually on a global basis, accounting for direct health care costs exceeding 6 billion dollars and making UTIs a very significant public health burden. This program project proposes studies that will improve our understanding of the epidemiology, microbial ecology, molecular pathogenesis and prevention of UTIs. The objectives of the project will be met through the development of a multidisciplinary research team that will collaborate to carry out four interrelated projects. Project 1 will undertake a double-blind placebo-controlled trial to determine whether a Lactobacillus crispatis vaginal probiotic will reduce the incidence of recurrent UTIs in women. The trial will also study adherence of the probiotic strain to vaginal epithelial cells from the subjects and will thus provide insight into the host and microbial mechanisms involved. Project 2 will address the hypothesis that cell surface glycosphingolipids in the bladder and vaginal epithelium are structurally organized into pleotrophic plasma membrane assemblies called caveolae and that caveolae participate in the initial epithelial response to attachment and uptake of uropathogenic e. coil Project 3 will employ two novel technologies that were developed during the previous funding period, namely high density microarrays and whole genome mutation scanning to characterize on a genomic level the molecular adaptation that uropathogenic E. coil strains undergo in the course of recurrent UTIs and in shifting from an asymptomatic to symptomatic infection. The project will also define how such adaptive evolution affects the ability of uropathogens to bind and invade epithelial cells, induce inflammation and resist phagocytosis. In Project 4, the association of host susceptibility to recurrent cystitis or pyelonephritis with known or novel mutations in specifically selected candidate host genes will be examined. The candidate genes to be studied are toll-like receptors (TLR2, TLR4, and TLR6), chemokine receptors (CXCRI and CXCR2), and interferon y receptors (IFN-yR1 and IFN-yR2). A laboratory core will provide microbiological studies, primary bladder and vaginal epithelial cells, characterization of urovirulence genes, bacterial adherence assays and other resources to the projects. An Administrative/Biostatistical Core will coordinate the overall project and provide biostatistical expertise to all investigators. The proposed studies will result in an improved understanding of pathogenetic mechanisms in UTIs and will result in new approaches to prevention of UTIs. KEY PERSONNEL, See instructions, Use continuation pages as needed to provide the required information Start with Principal Investigator, List all other key personnel in alphabetical order, last name first. Name Organization Walter E. Stamm, MD University of Washington Ann E. Stapleton, MD University of Washington Evgeni Sokurenko, MD, PhD University of Washington Thomas Mac Hooton, MD University of Washington Delia Scholes, PhD Center for Health Studies, Group Health Alan Aderem, PhD Institute for Systems Biology Lue Ping 7hao, PhD Fred Hutchinson Cancer Research Center Steve Moseley, PhD University of Washington Mark Stroud, PhD University of Washington Thomas Hawn, MD, PhD University of Washington/ISB Kalpana Gupta, MD, MPH University of Washington Sue Li, PhD Fred Hutchinson Cancer Research Center Continued, next page Disclosure Permission Statement, App!!cable to SBIR/STTR Only. See instructions. [] Yes in the format shown below, Role on Project Principal Investigator PI, Project 2 and Core B PI, Project 3 PI, Project 4 PI, GH Subcontract PI, ISB Subcontract PI, FHCRC Subcontract Co-Investigator Co-Investigator Co-Investigator Co-Investigator Co-Investigator o PHS 398 (Rev, 05/01) Page _2 Form Page 2 o [] Principal Investigator/ProgramDirector(Last,first, middle): Name, Continued Organization Roleon Project Harvard Medical School Consultant Anthony Atala, MD Sen-itiroh Hakomori, MD, PhD University of Washington Consultant Sharon Hillier, PhD " University of Pittsburgh Consultant Mary Claire King, PhD University of Washington Consultant Soman Abraham, PhD Duke University Consultant Richard Grady, MD University of Washington Consultant o PHS 398 (Rev. 05/01) Page 3 Form Page 3 o [] Principal Investigator/Program Director (Last, first, middle): Stamm, Walter E., M,D. PATHOGENIC MECHANISMS IN UTI NIDDK PROGRAM PROJECT GRANT APPLICATION RESEARCH GRANT TABLE OF CONTENTS Page Numbers A. Face Page .................................................................................................. 1 B. Description,